Materials that provide control over the release of multiple chemical or biological agents are of interest in a broad range of biomedical and biotechnological applications. [J. T. Santini, M. J. Cima, R. Langer, Nature 1999, 397, 335; L. D. Shea, E. Smiley, J. Bonadio, D. J. Mooney, Nat Biotechnol 1999, 17, 551; T. P. Richardson, M. C. Peters, A. B. Ennett, D. J. Mooney, Nat Biotechnol 2001, 19, 1029; W. M. Saltzman, W. L. Olbricht, Nat Rev Drug Discov 2002, 1, 177; A. C. R. Grayson, I. S. Choi, B. M. Tyler, P. P. Wang, H. Brem, M. J. Cima, R. Langer, Nat Mater 2003, 2, 767; J. M. Saul, M. P. Linnes, B. D. Ratner, C. M. Giachelli, S. H. Pun, Biomaterials 2007, 28, 4705.] Temporal control over the release of multiple biological cues, for example, will likely prove critical in applications such as tissue engineering, for which precise control over the administration of multiple different growth factors and other signals is thought to be required to promote the development of functional tissues. [Shea et al. 1999; Richardson et al. 2001; Saltzman et al. 2002; Saul et al. 2007.] Such sophisticated levels of control can also contribute to the development of new tools for basic biomedical research and more effective gene- and protein-based therapies. There is significant interest in the controlled release of anionic species, particularly anionic polypeptides and nucleic acids, including various forms of RNA and DNA.
Several recent reports have demonstrated approaches to the encapsulation of proteins or DNA in bulk matrices of degradable polymers or the fabrication of devices that provide control over the release of multiple agents [L. D. Shea, E. Smiley, J. Bonadio, D. J. Mooney, Nat Biotechnol 1999, 17, 551; T. P. Richardson, M. C. Peters, A. B. Ennett, D. J. Mooney, Nat Biotechnol 2001, 19, 1029; J. M. Saul, M. P. Linnes, B. D. Ratner, C. M. Giachelli, S. H. Pun, Biomaterials 2007, 28, 4705; Santini et al. 1999; Satlzman et al. 2002; Grayson et al. 2003.] Despite these advances, however, it has proven difficult to design thin films and coatings that provide control over the release of multiple proteins or DNA constructs with separate and distinct release profiles (e.g., rapid release of a first DNA construct, followed by the slower, sustained release of a second DNA construct). This invention relates generally to approaches to the fabrication of ultrathin polymer-based coatings that can be exploited to provide temporal control of release of anionic species. At least in part, the invention, relates to approaches to the controlled release of two or more anionic species with separate, distinct or both separate and distinct release profiles.
The present work relates to the use of methods developed for the layer-by-layer assembly of multilayered polyelectrolyte films (or ‘polyelectrolyte multilayers’). These methods are entirely aqueous and permit nanometer-scale control over the structures of thin films fabricated from a wide variety of synthetic or natural polyelectrolytes, including DNA. [G. Decher, Science 1997, 277, 1232; P. Bertrand, A. Jonas, A. Laschewsky, R. Legras, Macromol Rapid Comm 2000, 21, 319; P. T. Hammond, Adv Mater 2004, 16, 1271; Z. Y. Tang, Y. Wang, P. Podsiadlo, N. A. Kotov, Adv Mater 2006, 18, 3203; Y. Lvov, G. Decher, G. Sukhorukov, Macromolecules 1993, 26, 5396.]
Multilayers have been designed that release DNA and promote surface-mediated cell transfection by fabricating films using DNA and cationic polymers that are hydrolytically, enzymatically, or reductively degradable. Approaches to the fabrication, characterization, and application of DNA-containing multilayers have been reviewed recently. [D. M. Lynn, Soft Matter 2006, 2, 269; D. M. Lynn, Adv Mater 2007, 19, 4118.]
It has been reported that DNA can be incorporated into polyelectrolyte multilayers using layer-by-layer methods of assembly [Lvov, et al., 1993] and that it is possible to fabricate films that erode and release DNA in aqueous environments if the polycationic components of these assemblies are designed appropriately. [J. Zhang, L. S. Chua, D. M. Lynn, Langmuir 2004, 20, 8015; C. M. Jewell, J. Zhang, N. J. Fredin, D. M. Lynn, J. Control. Release. 2005, 106, 214; K. F. Ren, J. Ji, J. C. Shen, Biomaterials 2006, 27, 1152.; K. F. Ren, J. Ji, J. C. Shen, Bioconjugate Chem. 2006, 17, 77; C. M. Jewell, J. Zhang, N. J. Fredin, M. R. Wolff, T. A. Hacker, D. M. Lynn, Biomacromolecules 2006, 7, 2483; Blacklock, H. Handa, D. Soundara Manickam, G. Mao, A. Mukhopadhyay, D. Oupicky, Biomaterials 2007, 28, 117; J. Chen, S. Huang, W. Lin, R. Zhuo, Small 2007, 3, 636.] For example, it was recently reported that polyelectrolyte multilayers fabricated from plasmid DNA and hydrolytically degradable poly(beta-amino ester)s erode when incubated in physiological media [Zhang, et al. 2004 supra; Jewell, et al. 2005, supra; Jewell, et al. 2006, supra and D. M. Lynn, et al. 2006] and that objects coated with these assemblies promote surface-mediated transfection when placed in contact with mammalian cells. [Jewell et al. 2005, supra; Jewell et al. 2006, supra.]
It has also been reported that enzymatically or reductively degradable cationic polymers can be used to fabricate assemblies that release DNA in the presence of enzymes, reducing agents, or cells. [J. Zhang, et al. 2004; Jewell, et al. 2005; Zhang, et al. 2007; Ren, et al. Biomaterials 2006; K. F. Ren, J. Ji, J. C. Shen, Bioconjugate Chem. 2006, 17, 77; Blacklock, et al. 2007; Chen et al. 2007; N. Jessel, M. Oulad-Abdelghani, F. Meyer, P. Lavalle, Y. Haikel, P. Schaaf, J. C. Voegel, Proc Natl Acad Sci USA 2006, 103, 8618.]. These studies report approaches to promoting film erosion that involve the backbone degradation of cationic polymers and, in general, lead to films that release DNA relatively rapidly (e.g., over several hours to several days).
The present invention, in part, relates to an alternative approach to the disruption of ionic interactions in these assemblies that provides a means to extend the release of DNA or other anions over much longer periods (e.g., several months) particularly in ways that are useful in applications that require long-term exposure of cells or tissues to DNA. Additionally, the invention relates to approaches for the controlled release of two or more anions, particularly from a single multilayer film which exhibit rapid short-term release of one anion combined with long-term release of another anion.
U.S. Pat. No. 7,112,361 relates to decomposable films comprising a plurality of polyelectrolyte bilayers. Related published U.S. application 2007/0020469 reports decomposable films comprising a plurality of polyelectrolyte layers wherein a portion of the bilayers comprise a second entity selected from a biomolecule, a small molecule, a bioactive agent, and any combination thereof.
U.S. published application 20050027064 relates to charge-dynamic polymers useful for the delivery of anionic compounds including nucleic acids. The dynamic charge state cationic polymers are designed to have cationic charge densities that decrease by removal of removable functional groups from the polymers. The application also relates to complexes containing the polymers complexed to a polyanion and methods for using the interpolyelectrolyte complexes to deliver anionic compounds. The application describes compositions comprising a dynamic charge state cationic polymer, having a polymeric backbone formed from monomeric units, and having one or more removable functional groups attached to the polymeric backbone. The cationic charge of the dynamic charge state cationic polymer decreases when one or more of the removable functional groups is removed from the polymer. Specific dynamic charge state cationic polymers include those in which the polymer backbone comprises a polyamine, acrylate or methacrylate polymer, including polyethyleneimine, poly(propylene imine), poly(allyl amine), poly(vinyl amine), poly(amidoamine), or a dendrimer that is functionalized with terminal amine groups. The application also describes a method for delivering an anionic compound to a target cell by contacting a composition comprising a interpolyelectrolyte complex comprising a dynamic charge state cationic polymer complexed to one or more anions with the target cell thereby allowing the target cell to uptake the composition. After entry of the interpolyelectrolyte complex into the target cell, one or more of the removable functional groups is removed from the dynamic charge state cationic polymer decreasing the cationic charge of the dynamic charge state cationic polymer and promoting dissociation of the interpolyelectrolyte complex to release one or more anions.
U.S. published application 20060251701 relates to delivery of nucleic acids by polyelectrolyte assemblies formed by layer-by-layer deposition of nucleic acid and polycation and particularly to implantable medical devices coated with polyelectrolyte assemblies. Such devices facilitate the local delivery of a nucleic acid contained in the polyelectrolyte assembly into a cell or tissue at an implantation site.
The following references relate to formation of polyelectrolyte multilayers, dynamic charge state (charge shifting) polymers, release of anionic polyelectrolytes and/or drug release from thin films:
X. Liu, J. Zhang, and D. M. Lynn, “Polyelectrolyte Multilayers Fabricated from ‘Charge-Shifting’ Anionic Polymers: A New Approach to Controlled Film Disruption and the Release of Cationic Agents from Surfaces.” Soft Matter 2008, 4, 1688-1695; C. M. Jewell and D. M. Lynn, “Multilayered Polyelectrolyte Assemblies as Platforms for the Delivery of DNA and Other Nucleic Acid-Based Therapeutics.” Advanced Drug Delivery Reviews 2008, 60, 979-999; N. J. Fredin, J. Zhang, and D. M. Lynn, “Nanometer-Scale Decomposition of Ultrathin Multilayered Polyelectrolyte Films.” Langmuir 2007, 23, 2273-2276; J. Zhang, S. I. Montanez, C. M. Jewell, and D. M. Lynn, “Multilayered Films Fabricated from Plasmid DNA and a Side-Chain Functionalized Poly(beta-amino ester): Surface-Type Erosion and Sequential Release of Multiple Plasmid Constructs from Surfaces.” Langmuir 2007, 23, 11139-11146; J. Zhang and D. M. Lynn, “Ultrathin Multilayered Films Assembled from ‘Charge-Shifting’ Cationic Polymers: Extended, Long-Term Release of Plasmid DNA from Surfaces.” Advanced Materials 2007, 19, 4218-4223; D. M. Lynn, “Peeling Back the Layers: Controlled Erosion and Triggered Disassembly of Multilayered Polyelectrolyte Thin Films.” Advanced Materials 2007, 19, 4118-4130; J. Zhang, N. J. Fredin, J. F. Janz, B. Sun, and D. M. Lynn, “Structure/Property Relationships in Erodible Multilayered Films: Influence of Polycation Structure on Erosion Profiles and the Release of Anionic Polyelectrolytes.” Langmuir 2006, 22, 239-245; D. M. Lynn, “Layers of Opportunity: Nanostructured Polymer Assemblies for the Delivery of Macromolecular Therapeutics.” Soft Matter 2006, 2, 269-273; K. C. Wood, H. F. Chuang, R. D. Batten, D. M. Lynn, and P. T. Hammond, “Controlling Interlayer Diffusion to Achieve Sustained, Multi-Agent Delivery from Layer-by-Layer Films.” Proceedings of the National Academy of Sciences, USA 2006, 103, 10207-10212; J. Zhang, N. J. Fredin, and D. M. Lynn, “Erosion of Multilayered Assemblies Fabricated from Degradable Polyamines: Characterization and Evidence in Support of a Mechanism that Involves Polymer Hydrolysis.” Journal of Polymer Science—Part A: Polymer Chemistry 2006, 44, 5161-5173; C. M. Jewell, J. Zhang, N. J. Fredin, M. R. Wolff, T. A. Hacker, and D. M. Lynn, “Release of Plasmid DNA from Intravascular Stents Coated with Ultrathin Multilayered Polyelectrolyte Films.” Biomacromolecules 2006, 7, 2483-2491; J. Zhang and D. M. Lynn, “Multilayered Films Fabricated from Combinations of Degradable Polyamines Tunable Erosion and Release of Anionic Polyelectrolytes.” Macromolecules 2006, 39, 8928-8935.; C. M. Jewell, J. Zhang, N. J. Fredin, and D. M. Lynn, “Multilayered Polyelectrolyte Films Promote the Direct and Localized Delivery of DNA to Cells.” Journal of Controlled Release 2005, 106, 214-223; K. Wood, J. Q. Boedicker, D. M. Lynn, and P. T. Hammond, “Tunable Drug Release from Hydrolytically Degradable Layer-by-Layer Thin Films.” Langmuir 2005, 21, 1603-1609; N. J. Fredin, J. Zhang, and D. M. Lynn, “Surface Analysis of Erodible Multilayered Polyelectrolyte Films: Nanometer-Scale Structure and Erosion Profiles.” Langmuir 2005, 21, 5803-5811; and X. Liu, J. W. Yang, A. D. Miller, E. A. Nack, and D. M. Lynn, “Charge-Shifting Cationic Polymers that Promote Self-Assembly and Self-Disassembly with DNA.”Macromolecules 2005, 38, 7907-7914.
There is a need in the art for materials and methods that provide control over the release of multiple chemical or biological agents, particularly for controlled release of nucleic acids.